Proteomic Analysis of NF90 (Family) Complexes

描述(申请人提供)：NF90家族的双链RNA结合蛋白是由ILF3基因的转录本选择性剪接而产生的。这个家族的成员被发现在不同的亚细胞位置，并与不同的蛋白质复合体相互作用，提示在各种细胞过程中的功能，包括基因表达、DNA复制和修复、细胞生长和抗病毒免疫反应。虽然从cDNA中预测了至少7个独特的蛋白质产物，但已经确定在人类细胞系中占主导地位的是四个家族成员，并且这些亚型在瞬时表达分析中表现出不同程度的活性。这些观察结果增加了亚型参与特定复合体的可能性，这些复合体在细胞内显示出不同的功能。为了解决这一假设，含有NF90的天然复合体将从最近开发的、特征良好的表达四种异构体的稳定细胞系中分离出来。这些复合体的组成将使用蛋白质组学技术来确定。一旦知道了异步人类细胞的复杂成分(目标1)，将使用类似的方法来研究包含不同NF90成员的复合体作为细胞周期阶段和病毒感染的函数的动力学(目标2)。

DESCRIPTION (provided by applicant): The NF90 family of double-stranded RNA binding proteins is generated by alternative splicing of transcripts from the ILF3 gene. Members of this family have been found in several different subcellular locations and interact with varied protein complexes, suggestive of functions in various cellular processes including gene expression, DNA replication and repair, cell growth, and the anti-viral immune response. Although at least 7 unique protein products have been predicted from the cDNAs, it has been established that four family members predominate in human cell lines and that these isoforms exhibit different levels of activity in transient expression assays. These observations raise the possibility that the isoforms participate in specific complexes that display distinct functions within cells. To address this hypothesis, native NF90 containing complexes will be isolated from recently developed, well-characterized stable cell-lines expressing the four isoforms. The composition of the complexes will be established using proteomic techniques. Once the complex compositions are known for asynchronous human cells (Aim 1), a similar approach will be used to investigate the dynamics of the complexes containing different NF90 members as a function of cell cycle phase and viral infection (Aim 2).